Kidney transplantation is the only viable treatment for patients with end stage renal failure. However, the severe organ shortage leaves thousands in the US to suffer and die on waiting lists. Expanding the donor pool to include high risk donors may solve this shortage if those grafts could be preserved properly. However, organ preservation injury that is suffered after organ harvest is not well understood and little major advancement have been made in this area in the last 20 years. Therefore, the objective of this proposal is to identify novel molecular mechanisms of renal preservation injury involving the cytoskeleton system and to use this knowledge to direct future therapeutic approaches. Preliminary data show that the cytoskeleton protein ezrin fails during renal cold storage and that augmenting expression of ezrin significantly protects renal tubule epithelial cells from preservation injury. Conversely, ezrin knock-down does the opposite, which suggests causation. This proposal will test likely mechanisms of ezrin-induced cytoprotection in cell and kidney cold storage injury models by using molecular and genetic approaches to alter ezrin functionality. These studies will determine if ezrin, in its proper configuration, protects cell membrane structure and preserves cell mitochondrial function during cold ischemia and reperfusion. Renal epithelial cell models will be used where the cells express different amounts of wild type and mutant ezrin protein. Higher order models of preservation injury using isolated renal tubules and whole kidneys derived from transgenic ezrin knock-in mice expressing mutant ezrin genes will also be used. At the end of these studies, we will have gathered useful molecular information of how ezrin is involved in cold storage renal preservation injury and if this involvement is clinically relevant. These insights, together with previously identified mechanisms, may suggest likely therapeutic targets to improve renal preservation, which will allow harvesting of less robust kidneys from high risk donors to help alleviate the donor shortage. PUBLIC HEALTH RELEVANCE: Kidney transplantation is the treatment of choice for patients in end stage renal failure but there are not enough usable donor kidneys available. This need to expand the donor pool has increased the need to understand how to better preserve organs for transplantation, which in turn requires a more complete understanding of the mechanisms of organ preservation injury. This proposal is relevant to human health because it will bridge some of the key knowledge gaps in our understanding of the basic molecular mechanisms of how kidneys are injured after they are removed from the donor. This will reveal novel therapeutic targets to focus treatments and help alleviate the donor shortage.